Natural killer (NK) cells provide rapid early responses to viral infections and thus can contribute substantially to disease modulation and vaccine protection. Traditionally, NK cells have been considered to be nonspecific components of innate immunity, but recent studies in mice have shown that NK cells can also demonstrate features of antigen-specific memory. Until recently it was unclear whether this phenomenon might exist in primates, but our preliminary data demonstrates for the first time evidence of antigen-specific NK cell memory in any primate species. As compared with NK cells from uninfected macaques, splenic and hepatic NK cells from simian immunodeficiency virus (SIV)-infected animals were highly reactive to Gag- and Env-pulsed dendritic cells (DCs) but not to mock-pulsed DCs or mis-matched antigen controls. Similar Gag-specific NK cell responses were found in treated and untreated HIV-1- infected patients. Interestingly, memory NK cell responses in peripheral blood were low compared to distal sites, suggesting memory NK cells reside predominantly in tissues. We also found NK cell memory responses in rhesus macaques for over 5 years following vaccination with recombinant Ad26 vectors expressing HIV-1 Env or SIV Gag indicating antigen-specific NK cell memory is durable and does not require ongoing antigenic stimulation. The longevity, functionality, and specificity of memory NK cell responses in primates suggest their functional relevance and their potential importance against HIV-1 and other pathogens. In this new proposal we will address major deficits to this new field of study including identifying the mechanisms, kinetics, distribution, and efficacy of these responses using state-of-the-art techniques. We will evaluate the overarching hypothesis that memory NK cell responses elicited against SIV/HIV and adenovirus vaccine vectors are long-lived even in the absence of replicating virus and can be mobilized to engage and lyse virus-infected cells in an antigen- specific manner. We will evaluate this hypothesis with three focused Specific Aims: (1) Evaluate mobilization and recall of memory NK cells elicited against HIV and SIV vaccine antigens in humans and macaques; (2) Evaluate mobilization and maintenance of memory NK cells after SIV challenge in vaccinated and unvaccinated macaques; and (3) Explore cellular and molecular mechanisms of memory NK cell recognition and specificity.